Impairments in vascular endothelial function, particularly endothelium-dependent vasodilation, occurring in insulin resistant prediabetic states are thought to contribute to the accelerated rates of atherosclerotic vascular disease in type 2 diabetes. Endothelial vasodilatory dysfunction presents early in the pathogenesis of vascular disease, and contributes to the manifestation of atherogenic lesions, vasospasm, plaque rupture, intimal growth, and, in turn, coronary and cerebrovascular events. Moreover, forearm endothelial vasodilator dysfunction has been shown to be a marker of future cardiovascular events. Thus, a better understanding of the mechanisms responsible for the loss in endothelial vasodilator function associated with insulin resistance may lead to new targets for therapeutic intervention. Accordingly, the specific aims of the present proposal will be to determine: 1) if the blunted forearm endothelial vasodilator response to acetylcholine observed with obesity/insulin resistance reflects a specific agonist-related defect or rather a more general endothelial vasodilator abnormality; 2) whether the blunted forearm endothelial vasodilator response to acetylcholine observed with obesity/insulin resistance is related to: (a) decreased responsiveness to acetylcholine; (b) increase cholinesterase activity; (c) a selective impairment in stimulated nitric oxide release; (d) reduced muscarinic receptor function and/or number; and 3) if a program of regular endurance exercise improves endothelial vasodilator function, and whether the improvement is associated with increased insulin sensitivity. To address these aims, 180 middle-aged and older obese/insulin resistant and non-obese/insulin sensitive adults will be studied. Endothelium-dependent vasodilation will be assessed by changes in forearm blood flow (FBF: plethysmography) in response to intrabrachial infusions of acetylcholine, substance P, bradykinin, isoproterenol and methacholine. These endothelial agonists stimulate endothelial NO release via different cell surface receptors and intracellular G-protein-mediated signal transduction pathways. FBF responses to some agonists will also be determined in the presence of either NG.monomethyl arginine (nitric oxide synthase inhibitor) or atropine (muscarinic receptor blocker) to address specific aim 2c and 2d. Endothelial vasodilator function will also be assesed after a 3-month aerobic exercise program in a subgroup of obese/insulin resistant adults. The results of the proposed study should provide mechanistic insight into whether forearm endothelial vasodilator dysfunction in obese/insulin resistant adults is related to a specific receptor defect or a more general endothelial abnormality.